Presented to:

Presented by: Kevin Dischler, Bioassay Lab Director

Understanding Offshore Toxicity Testing – Beyond

“Pass” or “Fail”

2

Topics to be covered….

Drilling fluids – WBM and SBMProduced waterSubsea-chemicalsWhat are the limits?What do they mean?What makes a test “valid”?How are results derived?

Drilling Fluids – Toxicity Testing

Aquatic Toxicity – LC50 with Mysids (WBM/SBM) Tests the affect of the mud on the water

column LC50 = Lethal Concentration to 50% of the

exposed organisms Compliance target is an LC50 ≧ 30,000 ppm

SPP. SPP = Suspended Particulate Phase of 1 part

mud to 9 parts seawater mixture

4

Drilling Fluid LC50 Test Set-up

Control + 5 concentrations, 3 replicates, 20 shrimp per replicate, randomly loaded

Temp: 20±2ºC Salinity: 20±2 ppt 14 hours of light, 10 hours of dark SPP created by mixing mud and saltwater pH is adjusted

Test validity: Control must have at least 90% survival @ 96-

hours

Aquatic LC50Toxicity Test

Determining the LC50

% SPP Survivors (of 60)

Survival %

Control (0%) 59 98.3

20% 58 96.7

40% 54 90

60% 42 70

80% 24 40

100% 11 18.3

LC50 of this data set is 72.8% or 728,000 ppm SPPWith the target being ≧30,000 ppm, this would be a Passing test

Why a second drilling fluid tox test?

Sediment Toxicity Test

Measure the effect of the SBM coated cuttings on the sea floor

Uses Leptocheirus plumulosus as the test organism

Exposes the Leptos to a mixture of SBM and a formulated (man-made) sediment

Sediment Toxicity Test Set-up96-Hour tests are set-up with 5 concentrations

and a Control, 5 replicates, 20 Leptos per replicateTests are continuously aeratedTemp: 20±1ºCSalinity: 20±1 pptLight cycle: 14 hours of light, 10 hours of dark

Test validity: Check #1- Controls must have at least 90% survival @ 96-hours Check #2 - Controls must have Coefficient of Variation of <40%

With organisms from the same batch and randomly loaded:

A 96 Hour LC50 performed on the field sample

A 96 Hour LC50 performed on the appropriate reference mud:

- SBMs <11.0 ppg test with 9.0 ppg Ref Mud

- SBMs 11.0-14.0 ppg test with 11.5 ppg Ref Mud

- SBMs >14.0 ppg test vs. with 14.5 ppg Ref Mud

Sediment Toxicity Test Set-up

A SedTox test with a Ref Mud

- Randomly loaded organisms- Randomly placed on the test shelves

The Sediment Toxicity Ratio (STR)

The compliance limit for an STR is ≦ 1.0

LC50 Value for the Reference Drilling Fluid = STR

LC50 Value for the Submitted SBM sample  

Ex: Ref MudLC50 = 42.9 ml/Kg

Field Mud LC50 = 197 ml/Kg

42.9 ÷ 197 = 0.2 Sediment Toxicity Ratio is 0.2 and is a Pass

But what if…..? SedTox can be a difficult test. So much so,

that EPA allows it to be run three times.

An STR of >1.0 on the first test is an “initial failure.” Not out of compliance yet.

Test 2 must be run from same sample, same bottle.

Test 3 must be run from a sample caught within 15 minutes of the first sample

When averaging is needed, the LC50s are averaged, not the STRs.

14

Produced Water Toxicity Test Chronic static renewal seven

(7) day survival and growth test

Measures the effect of Produced Water on the survival and growth of the test organisms.

The Vertebrate organism is a Menidia beryllina

The Invertebrate organism is Mysidopsis bahia

15

Produced Water Toxicity Test Set-up

Mysid portion: - 7 day old organisms - Control + 5 concentrations, 8 replicates of 5 shrimp - Temperature: 26±1ºC, Salinity: 25±2 ppt Menidia (minnow) portion: - 7 to 11 day old organisms - Control + 5 concentrations, 5 replicates of 8 fish - Temperature: 25±1ºC, Salinity: 25±2 ppt Both require: - 16 hours of light, 8 hours of dark - 0.5 Dilution series – concentrations halve and double

16

Produced Water Toxicity TestRandomly loaded organisms

Randomly placed on test shelves

This is a “renewal” test. Test water carefully changed out daily, survivors counted and recorded daily.

17

Looking for a “Passing” NOEC

NOEC is No Observed Effect Concentration

A NOEC for survival < Critical Dilution is considered a

lethal effect and requires re-testing

A NOEC for growth < Critical Dilution is considered a sub- lethal effect and requires re-testing

NOEC > Critical Dilution required to pass

18

What is a Critical Dilution?The Critical Dilution is determined by three factors used

in Table 1 Appendix D from the GMG Permit

- Highest dailly average flow rate for the previous 3 months prior to sampling- Diameter of the discharge pipe- Water depth from discharge pipe to seafloor

What does the Critical Dilution represent? The Cormix model tells us that at a point 100 m from

the outfall, a random sample pulled from the sea will contain that same % of effluent from the outfall.

Produced Water Toxicity Test

At test completion, survival counts are recorded.

The surviving organisms are dried overnight, cooled and then weighed

Produced Water Toxicity Test Validity

Check #1: Controls must have a minimum of 80% survival @ 7 days

Check #2: Mysids in Control must average 0.20 mg

Check #3: Minnows in Control must average 0.50 mg

Check #4: Coefficient of Variation for the Controls must be <40%

Check #5: Unless significant effects are exhibited, the Coefficient of Variation for Critical Dilutions must be <40%

Check #6: A Percent Minimum Significant Difference range of 11-37 for Mysid growth and 11-28 for Menidia growth shall be applied accordingly.

A test failure can not be deemed invalid due to a CV exceedence

Time of Reading Control (0%) 0.04% 0.08% 0.16% 0.32% 0.64%

24 hours 97.5 97.5 100 100 97.5 100

48 hours 95 97.5 100 100 95 100

7 days 87.5 95 95 95 82.5 95

CV% 17.6 8.57 8.57 8.57 16.8 8.57

DATA TABLE FOR Mysidopsis bahia SURVIVAL

Time of Reading Control (0%) 0.04% 0.08% 0.16% 0.32% 0.64%

24 hours 97.5 97.5 100 100 97.5 100

48 hours 90 90 100 90 87.5 80

7 days 77.5 85 87.5 72.5 65 55

CV% 17.6 8.57 8.57 16.6 16.8 8.57

DATA TABLE FOR Mysidopsis bahia GROWTH

% A B C D E CV%0 0.770 0.699 0.753 0.719 0.649 0.718 6.6

0.04 0.727 0.683 0.794 0.727 0.716 0.729 5.50.08 0.794 0.712 0.723 0.67 0.716 0.723 6.20.16 0.808 0.362 0.225 0.959 0.912 0.653 51.50.32 0.659 0.764 0.668 0.809 0.651 0.710 10.10.64 0.491 0.651 0.916 0.618 0.747 0.685 24.8

Average dry weight (mg) Mean Dry Wt.

% A B C D E CV%0 0.770 0.699 0.753 0.719 0.649 0.718 6.6

0.04 0.707 0.683 0.651 0.615 0.695 0.670 5.50.08 0.711 0.715 0.695 0.654 0.671 0.689 6.20.16 0.699 0.689 0.691 0.651 0.689 0.684 15.50.32 0.612 0.595 0.562 0.511 0.503 0.557* 10.10.64 0.491 0.412 0.395 0.396 0.405 0.421* 24.8

Average dry weight (mg) Mean Dry Wt.

Sub-sea Fluid Toxicity testingExamples: Sub-sea wellhead preservation fluids,

leak tracer fluids, umbilical storage fluids, riser tensioner fluids, etc.

The permit sets the limit or Critical Dilution at 50 mg/l

Same method as Produced Water Toxicity Test

Same organisms and test criteria

24

“Partners in Compliance”

On December 16th, 2013

>>>>>>>>>